Glyphosate, the active ingredient in RoundUp produced by Monsanto, is the most commonly used pesticides in the US across agricultural, industrial, and home settings with 180-185 million pounds used in the US in 2007. \ While glyphosate exposure has been linked to adverse health effects, there is a surprising lack of mechanistic information concerning the full scope of long-term health effects and potential toxicities associated with glyphosate exposure. Understanding how glyphosate interacts with biological systems in vivo in mammals is absolutely necessary to assess the prolonged effects and mechanism of toxicity of glyphosate on human health. This project will apply innovative chemical technologies to map the direct proteome-wide targets of glyphosate and its potentially reactive metabolites in vivo, as well as investigate the effects co-exposures to glyphosate and chemical mixtures that act through overlapping mechanisms. Here, we will use an innovative chemoproteomic strategy termed reactivity-based protein profiling (RBPP), which uses reactivity-based chemical probes to identify hyper-reactive protein hotspots in complex proteomes, to map direct proteome-wide targets of glyphosate. Using the RBPP platform, we have found that glyphosate as well as several high usage environmental chemicals of concern commonly inhibit several metabolic enzymes involved in fatty acid degradation and metabolism. We hypothesize that cumulative exposure to glyphosate and other environmental chemicals that commonly inhibit fatty acid degradation enzymes will directly cause additive or synergistic lipid dysregulation, tissue adiposity, and serum dyslipidemia in vivo in mice. We propose to apply innovative chemoproteomic platforms to map proteome-wide targets of glyphosate to reveal novel toxicological mechanisms of this widely used and controversial herbicide, with a particular focus on understanding how exposure to glyphosate and other chemical and nutritional exposures may synergize to impact fat metabolism.